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Frontiers in Microbiology 2018is a bacterial plant pathogen that causes soft rot disease on a wide range of host plants. The type III secretion system (T3SS) is an important virulence factor in ....
Development of a Method to Monitor Gene Expression in Single Bacterial Cells During the Interaction With Plants and Use to Study the Expression of the Type III Secretion System in Single Cells of in Potato.
is a bacterial plant pathogen that causes soft rot disease on a wide range of host plants. The type III secretion system (T3SS) is an important virulence factor in . Expression of the T3SS is induced in the plant apoplast or in -inducing minimal medium (hrp-MM), and is repressed in nutrient-rich media. Despite the understanding of induction conditions, how individual cells in a clonal bacterial population respond to these conditions and modulate T3SS expression is not well understood. In our previous study, we reported that in a clonal population, only a small proportion of bacteria highly expressed T3SS genes while the majority of the population did not express T3SS genes under hrp-MM condition. In this study, we developed a method that enabled observation and quantification of gene expression in single bacterial cells . Using this technique, we observed that the expression of the T3SS genes and is restricted to a small proportion of cells during the infection of potato. We also report that the expression of T3SS genes is higher at early stages of infection compared to later stages. This expression modulation is achieved through adjusting the ratio of T3SS and T3SS cells and the expression intensity of T3SS cells. Our findings not only shed light into how bacteria use a bi-stable gene expression manner to modulate an important virulence factor, but also provide a useful tool to study gene expression in individual bacterial cells .
PubMed: 30002651
DOI: 10.3389/fmicb.2018.01429 -
International Journal of Molecular... May 2021Type II toxin-antitoxin (TA) systems are genetic elements usually encoding two proteins: a stable toxin and an antitoxin, which binds the toxin and neutralizes its toxic...
Type II toxin-antitoxin (TA) systems are genetic elements usually encoding two proteins: a stable toxin and an antitoxin, which binds the toxin and neutralizes its toxic effect. The disturbance in the intracellular toxin and antitoxin ratio typically leads to inhibition of bacterial growth or bacterial cell death. Despite the fact that TA modules are widespread in bacteria and archaea, the biological role of these systems is ambiguous. Nevertheless, a number of studies suggests that the TA modules are engaged in such important processes as biofilm formation, stress response or virulence and maintenance of mobile genetic elements. The 3937 strain serves as a model for pathogens causing the soft-rot disease in a wide range of angiosperm plants. Until now, several chromosome-encoded type II TA systems were identified in silico in the genome of this economically important bacterium however so far only one of them was experimentally validated. In this study, we investigated three putative type II TA systems in 3937: , and , which represents a novel toxin/antitoxin superfamily. We provide an experimental proof for their functionality in vivo both in and . Finally, we examined the prevalence of those systems across the Pectobacteriaceae family by a phylogenetic analysis.
Topics: Bacterial Proteins; Bacterial Toxins; Dickeya; Gene Expression Regulation, Bacterial; Plant Diseases; Toxin-Antitoxin Systems; Virulence
PubMed: 34073004
DOI: 10.3390/ijms22115932 -
Infection and Immunity May 2017Type II secretion (T2S) is one means by which Gram-negative pathogens secrete proteins into the extracellular milieu and/or host organisms. Based upon recent genome... (Review)
Review
Type II secretion (T2S) is one means by which Gram-negative pathogens secrete proteins into the extracellular milieu and/or host organisms. Based upon recent genome sequencing, it is clear that T2S is largely restricted to the , occurring in many, but not all, genera in the , , , and classes. Prominent human and/or animal pathogens that express a T2S system(s) include , , , , , , , , , and T2S-expressing plant pathogens include , , , , , , and T2S also occurs in nonpathogenic bacteria, facilitating symbioses, among other things. The output of a T2S system can range from only one to dozens of secreted proteins, encompassing a diverse array of toxins, degradative enzymes, and other effectors, including novel proteins. Pathogenic processes mediated by T2S include the death of host cells, degradation of tissue, suppression of innate immunity, adherence to host surfaces, biofilm formation, invasion into and growth within host cells, nutrient assimilation, and alterations in host ion flux. The reach of T2S is perhaps best illustrated by those bacteria that clearly use it for both environmental survival and virulence; e.g., employs T2S for infection of amoebae, growth within lung cells, dampening of cytokines, and tissue destruction. This minireview provides an update on the types of bacteria that have T2S, the kinds of proteins that are secreted via T2S, and how T2S substrates promote infection.
Topics: Bacterial Proteins; Gram-Negative Bacteria; Protein Transport; Type II Secretion Systems; Virulence Factors
PubMed: 28264910
DOI: 10.1128/IAI.00014-17 -
Innovation and Application of the Type III Secretion System Inhibitors in Plant Pathogenic Bacteria.Microorganisms Dec 2020Many Gram-negative pathogenic bacteria rely on a functional type III secretion system (T3SS), which injects multiple effector proteins into eukaryotic host cells, for... (Review)
Review
Many Gram-negative pathogenic bacteria rely on a functional type III secretion system (T3SS), which injects multiple effector proteins into eukaryotic host cells, for their pathogenicity. Genetic studies conducted in different host-microbe pathosystems often revealed a sophisticated regulatory mechanism of their T3SSs, suggesting that the expression of T3SS is tightly controlled and constantly monitored by bacteria in response to the ever-changing host environment. Therefore, it is critical to understand the regulation of T3SS in pathogenic bacteria for successful disease management. This review focuses on a model plant pathogen, , and summarizes the current knowledge of its T3SS regulation. We highlight the roles of several T3SS regulators that were recently discovered, including the transcriptional regulators: FlhDC, RpoS, and SlyA; the post-transcriptional regulators: PNPase, Hfq with its dependent sRNA ArcZ, and the RsmA/B system; and the bacterial second messenger cyclic-di-GMP (c-di-GMP). Homologs of these regulatory components have also been characterized in almost all major bacterial plant pathogens like , , spp., spp., and spp. The second half of this review shifts focus to an in-depth discussion of the innovation and development of T3SS inhibitors, small molecules that inhibit T3SSs, in the field of plant pathology. This includes T3SS inhibitors that are derived from plant phenolic compounds, plant coumarins, and salicylidene acylhydrazides. We also discuss their modes of action in bacteria and application for controlling plant diseases.
PubMed: 33317075
DOI: 10.3390/microorganisms8121956 -
Frontiers in Plant Science 2017The production of reactive oxygen species (ROS) is one of the first defense reactions induced in in response to infection by the pectinolytic enterobacterium . Previous...
The production of reactive oxygen species (ROS) is one of the first defense reactions induced in in response to infection by the pectinolytic enterobacterium . Previous results also suggest that abscisic acid (ABA) favors multiplication and spread into its hosts. Here, we confirm this hypothesis using ABA-deficient and ABA-overproducer plants. We investigated the relationships between ABA status and ROS production in after infection and showed that ABA status modulates the capacity of the plant to produce ROS in response to infection by decreasing the production of class III peroxidases. This mechanism takes place independently of the well-described oxidative stress related to the RBOHD NADPH oxidase. In addition to this weakening of plant defense, ABA content in the plant correlates positively with the production of some bacterial virulence factors during the first stages of infection. Both processes should enhance disease progression in presence of high ABA content. Given that infection increases transcript abundance for the ABA biosynthesis genes and and triggers ABA accumulation in leaves, we propose that manipulates ABA homeostasis as part of its virulence strategy.
PubMed: 28421092
DOI: 10.3389/fpls.2017.00456 -
Research in Microbiology 2015We previously showed that SlyA of Dickeya dadantii 3937 plays an important role in virulence toward plants, and that the ΔslyA mutant is hypermotile, whereas flagellum...
We previously showed that SlyA of Dickeya dadantii 3937 plays an important role in virulence toward plants, and that the ΔslyA mutant is hypermotile, whereas flagellum synthesis and flagellin production are indistinguishable from the wild type. Here we show that motility factors, including the distance of continuous directed movement, time for that movement and speed, were significantly higher in the ΔslyA mutant than in the wild type. Remarkably, transcription levels of motA and motB, that are involved in flagellar rotation, were elevated in the ΔslyA mutant, suggesting that the mutant's hypermotility was due to an increase in flagellar rotation. In low (10 μM) magnesium medium that activates the PhoP-PhoQ system, growth and virulence of the ΔslyA mutant were much lower than for the wild type; expression of motA, motB, mgtA, pelA, pelB, pelC, pelD, pelE, pelI, indA, tolC, sodC, acsA and hrpN were also reduced in the mutant. Interestingly, motA, motB, pelD, pelE, pelI, sodC and indA were also reduced in phoP and phoQ mutants. Because the SlyA protein directly binds to the promoter region of PhoP, SlyA regulates virulence by controlling multiple pathogenicity-related genes directly and/or at least by controlling PhoP in D. dadantii 3937 when magnesium is low.
Topics: Bacterial Proteins; Enterobacteriaceae; Flagella; Gene Expression Regulation, Bacterial; Magnesium Sulfate; Mutation; Stress, Physiological; Transcription, Genetic; Virulence
PubMed: 26027774
DOI: 10.1016/j.resmic.2015.05.004 -
Environmental Microbiology Nov 2016Dickeya species are soft rot disease-causing bacterial plant pathogens and an emerging agricultural threat in Europe. Environmental modulation of gene expression is...
Dickeya species are soft rot disease-causing bacterial plant pathogens and an emerging agricultural threat in Europe. Environmental modulation of gene expression is critical for Dickeya dadantii pathogenesis. While the bacterium uses various environmental cues to distinguish between its habitats, an intricate transcriptional control system coordinating the expression of virulence genes ensures efficient infection. Understanding of this behaviour requires a detailed knowledge of expression patterns under a wide range of environmental conditions, which is currently lacking. To obtain a comprehensive picture of this adaptive response, we devised a strategy to examine the D. dadantii transcriptome in a series of 32 infection-relevant conditions encountered in the hosts. We propose a temporal map of the bacterial response to various stress conditions and show that D. dadantii elicits complex genetic behaviour combining common stress-response genes with distinct sets of genes specifically induced under each particular stress. Comparison of our dataset with an in planta expression profile reveals the combined impact of stress factors and enables us to predict the major stress confronting D. dadantii at a particular stage of infection. We provide a comprehensive catalog of D. dadantii genomic responses to environmentally relevant stimuli, thus facilitating future studies of this important plant pathogen.
Topics: Bacterial Proteins; Enterobacteriaceae; Europe; Gene Expression Regulation, Bacterial; Genomics; Plant Diseases; Plants; Virulence; Virulence Factors
PubMed: 26940633
DOI: 10.1111/1462-2920.13267 -
Applied and Environmental Microbiology Nov 2016Modification of teichoic acid through the incorporation of d-alanine confers resistance in Gram-positive bacteria to antimicrobial peptides (AMPs). This process involves...
UNLABELLED
Modification of teichoic acid through the incorporation of d-alanine confers resistance in Gram-positive bacteria to antimicrobial peptides (AMPs). This process involves the products of the dltXABCD genes. These genes are widespread in Gram-positive bacteria, and they are also found in a few Gram-negative bacteria. Notably, these genes are present in all soft-rot enterobacteria (Pectobacterium and Dickeya) whose dltDXBAC operons have been sequenced. We studied the function and regulation of these genes in Dickeya dadantii dltB expression was induced in the presence of the AMP polymyxin. It was not regulated by PhoP, which controls the expression of some genes involved in AMP resistance, but was regulated by ArcA, which has been identified as an activator of genes involved in AMP resistance. However, arcA was not the regulator responsible for polymyxin induction of these genes in this bacterium, which underlines the complexity of the mechanisms controlling AMP resistance in D. dadantii Two other genes involved in resistance to AMPs have also been characterized, phoS and phoH dltB, phoS, phoH, and arcA but not dltD mutants were more sensitive to polymyxin than the wild-type strain. Decreased fitness of the dltB, phoS, and phoH mutants in chicory leaves indicates that their products are important for resistance to plant AMPs.
IMPORTANCE
Gram-negative bacteria can modify their lipopolysaccharides (LPSs) to resist antimicrobial peptides (AMPs). Soft-rot enterobacteria (Dickeya and Pectobacterium spp.) possess homologues of the dlt genes in their genomes which, in Gram-positive bacteria, are involved in resistance to AMPs. In this study, we show that these genes confer resistance to AMPs, probably by modifying LPSs, and that they are required for the fitness of the bacteria during plant infection. Two other new genes involved in resistance were also analyzed. These results show that bacterial resistance to AMPs can occur in bacteria through many different mechanisms that need to be characterized.
Topics: Anti-Bacterial Agents; Antimicrobial Cationic Peptides; Bacterial Proteins; Cichorium intybus; Drug Resistance, Bacterial; Enterobacteriaceae; Gene Expression Regulation, Bacterial; Mutation; Plant Leaves; Polymyxins; Repressor Proteins
PubMed: 27565623
DOI: 10.1128/AEM.01757-16 -
Nucleic Acids Research Sep 2022DNA supercoiling is an essential mechanism of bacterial chromosome compaction, whose level is mainly regulated by topoisomerase I and DNA gyrase. Inhibiting either of...
DNA supercoiling is an essential mechanism of bacterial chromosome compaction, whose level is mainly regulated by topoisomerase I and DNA gyrase. Inhibiting either of these enzymes with antibiotics leads to global supercoiling modifications and subsequent changes in global gene expression. In previous studies, genes responding to DNA relaxation induced by DNA gyrase inhibition were categorised as 'supercoiling-sensitive'. Here, we studied the opposite variation of DNA supercoiling in the phytopathogen Dickeya dadantii using the non-marketed antibiotic seconeolitsine. We showed that the drug is active against topoisomerase I from this species, and analysed the first transcriptomic response of a Gram-negative bacterium to topoisomerase I inhibition. We find that the responding genes essentially differ from those observed after DNA relaxation, and further depend on the growth phase. We characterised these genes at the functional level, and also detected distinct patterns in terms of expression level, spatial and orientational organisation along the chromosome. Altogether, these results highlight that the supercoiling-sensitivity is a complex feature, which depends on the action of specific topoisomerases, on the physiological conditions, and on their genomic context. Based on previous in vitro expression data of several promoters, we propose a qualitative model of SC-dependent regulation that accounts for many of the contrasting transcriptomic features observed after DNA gyrase or topoisomerase I inhibition.
Topics: DNA Gyrase; DNA Topoisomerases, Type I; DNA, Superhelical; DNA, Bacterial; Enterobacteriaceae; Anti-Bacterial Agents
PubMed: 35950487
DOI: 10.1093/nar/gkac679 -
The Journal of Biological Chemistry Dec 2022Succination is the spontaneous reaction between the respiratory intermediate fumarate and cellular thiols that forms stable S-(2-succino)-adducts such as...
Succination is the spontaneous reaction between the respiratory intermediate fumarate and cellular thiols that forms stable S-(2-succino)-adducts such as S-(2-succino)cysteine (2SC). 2SC is a biomarker for conditions associated with elevated fumarate levels, including diabetes, obesity, and certain cancers, and succination likely contributes to disease progression. Bacillus subtilis has a yxe operon-encoded breakdown pathway for 2SC that involves three distinct enzymatic conversions. The first step is N-acetylation of 2SC by YxeL to form N-acetyl-2SC (2SNAC). YxeK catalyzes the oxygenation of 2SNAC, resulting in its breakdown to oxaloacetate and N-acetylcysteine, which is deacetylated by YxeP to give cysteine. The monooxygenase YxeK is key to the pathway but is rare, with close homologs occurring infrequently in prokaryote and fungal genomes. The existence of additional 2SC breakdown pathways was not known prior to this study. Here, we used comparative genomics to identify a S-(2-succino) lyase (2SL) that replaces yxeK in some yxe gene clusters. 2SL genes from Enterococcus italicus and Dickeya dadantii complement B. subtilis yxeK mutants. We also determined that recombinant 2SL enzymes efficiently break down 2SNAC into fumarate and N-acetylcysteine, can perform the reverse reaction, and have minor activity against 2SC and other small molecule thiols. The strong preferences both YxeK and 2SL enzymes have for 2SNAC indicate that 2SC acetylation is a conserved breakdown step. The identification of a second naturally occurring 2SC breakdown pathway underscores the importance of 2SC catabolism and defines a general strategy for 2SC breakdown involving acetylation, breakdown, and deacetylation.
Topics: Cysteine; Lyases; Acetylcysteine; Sulfhydryl Compounds; Fumarates
PubMed: 36309089
DOI: 10.1016/j.jbc.2022.102639